During the morphogenesis of several animal and bacterial DNA viruses, DNA is packaged in a proteinaceous envelope. Our long-range objective is to understand the mechanism of viral DNA packaging at the molecular level. To reach this goal, we will use bacteriophage T7 as a model system and will pursue the following short-range objectives: a) isolation of intermediates in the T7 DNA packaging pathway. Lysates of T7-infected E. Coli and cell-free extracts that package DNA "in vitro" will be fractionated. Mutational and chemical alteration of the DNA packaging pathway will be used to isolate intermediates that are transient or unstable during normal T7 assembly. To assist in determining the temporal order of DNA packaging intermediates, kinetic labeling experiments will be performed. The ability of isolated intermediates to participate in DNA packaging in vitro will be tested. b) Characterization of DNA packaging intermediates. To determine the mechanisms by which intermediates progress in the T7 DNA packaging pathway, we will characterize the intermediates isolated. The techniques to be used for characterization include: buoyant density and velocity ultracentrifugation, electron microscopy, electrophoresis (non-denaturing) in agarose gels and density gradients, sodium dodecyl sulfate polyacrylamide gel electrophoresis and quasielastic laser light scattering. c) Continuation of the development of improved techniques for isolating and characterizing DNA packaging intermediates. These improved techniques will be used in a) and b). Emphasis will be placed on improving existing techniques for electrophoresis, developing techniques for partitioning in aqueous-polymer multiphase systems and improving techniques to prepare intermediates for electron microscopy. d) Development of dynamic models of the DNA packaging process from the above data and the testing of these models.